In the vertebrate nervous system, dopaminergic (DA) neurons are detected in small clusters and at stereotypic locations. The importance of these neurons is underscored by their involvement in multiple human neurological disorders including Parkinson's disease, schizophrenia, addiction, and autism. Elucidation of the mechanisms that determine the identity and connectivity of DA neurons is essential to defining and interpreting the causes of these disorders. Meanwhile, understanding the development of DA neurons will shed important light on fundamental mechanisms governing processes ranging from in vivo progenitor cell behavior to neural circuit formation in the vertebrate nervous system. This application proposes to continue research into the development of DA neurons, using zebrafish as a genetic model organism. In the past five years, our functional study of DA system suggests that the ventral forebrain DA neurons in zebrafish regulate movement, reward-associated behaviors, and neuroendocrine function. Our developmental study of DA system identifies the evolutionarily conserved transcription regulator Neurogenin 1 (Ngn1) as an important determinant of these DA neurons, and the conserved zinc finger protein Tof/Fezl, as an upstream regulator of Ngn1. Moreover, we established a transgenic line in which DA neurons are labeled with GFP reporter, allowing their visualization in living embryos and larvae. In this proposal, we wish to build upon these findings, to address the following questions: 1) What are the cellular mechanisms by which tof/fezl and ngn1 regulate the commitment of progenitor cells to DA neurons? 2) What are the molecular relationships between tof/fezl and ngn1 in DA neuron development? 3) What are the downstream effectors of tof/fezl and ngn1 in DA neuron development? The proposed studies are likely to have a long-term impact on understanding in vivo stem cell behavior and developing replacement or regenerative therapies, by advancing our understanding of the mechanisms underlying neural progenitor commitment and differentiation into DA neurons. PUBLIC HEALTH REVELANCE: Our research aims to understand the production of dopaminergic (DA) neurons using zebrafish, a powerful genetic model organism with transparent embryonic and larval stages. The importance of DA neurons is underscored by their involvement in multiple human neurological disorders including Parkinson's disease, schizophrenia, addiction and autism. Our proposed studies are likely to have a long-term impact on understanding in vivo stem cell behavior and developing replacement or regenerative therapies to treat neurological disorders such as Parkinson's disease.